The present invention relates to optically pure S-(xe2x88x92)-benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof, substantially free of their R-(+)-isomers, to processes for preparation of the optically pure S-(xe2x88x92)-benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof substantially free of their R-(+)-isomers, and to pharmaceutical compositions comprising the S(xe2x88x92)-benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof. These compounds and compositions can be used to systemically and topically treat bacterial Gram-positive, Gram-negative and anaerobic infections, specially resistant Gram-positive organism infections, Gram-negative organism infections, mycobacterial infections and emerging nosocomial pathogen infections, while avoiding toxic effects associated with the administration of the racemic mixture of RS-(xc2x1)-benzoquinolizine carboxylic acid. The compounds and compositions of this invention can also be used to treat diseases and disorders caused by Gram-positive, Gram-negative and anaerobic bacteria, and diseases and disorders caused by resistant Gram-positive organisms, Gram-negative organisms, mycobacteria and nosocomial pathogens.
Bacterial resistance to antibiotics is an increasingly recurrent phenomenon. Of grave concern has been the development of methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant Streptococcus epidermidis (MRSE) strains, which because of the phenomenon of cross-resistance, are now also resistant to the larger class of xcex2-lactam antibiotics including the cephalosporins and carbapenems. Of even graver concern is the development of resistance in MRSA strains against the class of anti-bacterial agents known as fluoroquinolones. Several reports are known of MRSA strains displaying resistance to fluoroquinolone agents such as ciprofloxacin, sparfloxacin and even the more recently introduced trovafloxacin. In addition, for trovafloxacin and for newer introductions like grepafloxacin, moxifloxacin and gatifloxacin, a concern has been expressed about their checkered safety records. The use of trovafloxacin has been suspended or severely curtailed because of its association with liver side effects. Grepafloxacin was withdrawn worldwide because of severe cardiovascular side effects. The labelling on gatifloxacin and moxifloxacin warns that they may prolong the QTc interval on electrocardiograms in some patients.
The last line of defense against such fluoroquinolone-resistant MRSA strains is the class of glycopeptide antibiotics represented by vancomycin and teicoplanin. These glycopeptide antibiotics are, however, laden with several limitations. Vancomycin is encumbered with lack of oral bioavailability, nephrotoxic potential, toxic effects such as phlebitis and red-men syndrome. Moreover, the recent disturbing wide spread emergence of Vancomycin resistant enterococci (VRE) followed by the alarming reports of Vancomycin intermediate resistance Staphylococcus aureus (VISA) strains from Japan and USA have cast a shadow over the future of glycoside antibiotics in clinical practice. In time, there is a relatively wide-spread emergence of staphylococci, enterococci, pneumococci and streptococci, which have become resistant to currently used first-and second-line antibacterial agents such as penicillin, oxacillin, vancomycin and erythromycin (SENTRY Programme: Antimicrobial Agents and Chemotherapy 42 1762-1770, 1998).
Also, for primary skin infections such as impetigo and folliculitis, and for secondary infections in humans such as infected dermatitis, wounds and bums, as well as to eliminate nasal carriage of MRSA in healthcare workers and patients, a special antibiotic used topically is Mupirocin. Mupirocin has high in vitro anti-staphylococcal and anti-streptococcal activity. There has, however, been an increase of organisms, specially staphylococci, developing resistance to Mupirocin. The emergence of Mupirocin-resistant Methicillin-resistant Staphylococcus aureus (MRSA) in infected patients in different countries like Canada, Western Australia, UK, Spain and Switzerland is described in different references in the medical and scientific literature viz. J. Hosp. Infect. 39(1), 19-26 (1998); J. Hosp. Infect. 26(3),157-165 (1994); Infect Control Epidemiol 17(2), 811-813 (1996); 38th Annual ICAAC Abstract C-75, 90 (1998); 38th ICAAC Abstract 12-25, 507 (1998).
Furthermore, Gram-positive pathogens such as Staphylococci, enterococci and Gram-negative pathogens E. coli, Klebsiella, Proteus, Serratia, Citrobacter and Pseudomonas, frequently encountered in urinary tract infections are susceptible to the known fluoroquinolones, such as ciprofloxacin, levofloxacin, ofloxacin and norfloxacin. The potency of these fluoroquinolones, however, markedly deteriorates under the acidic conditions likely to be encountered in urinary tract infections, rendering them inadequate.
Furthermore, multidrug-reistant (MDR) mycobacterial strains have emerged displaying resistance to first-line antimycobacterial agents such as rifampicin, pyrazinamide and INH etc. thus severely curtailing therapeutic options available for the management of infections due to such strains. Usually, the antimycobacterial drug regimen involves treatment spread over several months, and hence the drug has to be tolerated well by the patients. Among the fluoroquinolone antibiotics, sparfloxacin is reported to be highly active against mycobacteria. It is not quite suitable, however, for long-term therapy because of its potential to cause phototoxic side effects in humans and laboratory animals such as mice and guinea-pigs.
Furthermore, in the worldwide management of nosocomial infections, besides the problematic strains of staphylococci and enterococci, including MRSA, strains of Chryseobacteria have recently emerged as new members of nosocomial pathogens causing neonatal meningitis and pneumonia, as well as sepsis, in immuno-compromised patients being treated in intensive care units. Chryseobacteria are intrinsically resistant to xcex2-lactam antibiotics including third-generation cephalosporins and carbapenems. These factors reduce the treatment options available to the clinicians.
The highly pressing need for other agents and methods of treatment for infections arising from such emerging resistant microorganisms, Gram-negative pathogens in acidic environments, mycobacteria and nosocomial pathogens thus assumes great significance.
Among other agents, one particular class of compounds the benzoquinolizine carboxylic acids are of particular relevance. Nadifloxacin is an example of a benzo-quinolizine carboxylic acid. Nadifloxacin is racemic [(xc2x1)-9-fluoro-8-(4-hydroxypiperidin-1-yl)-5-methyl-6,7-dihydro-1-oxo-1H-5H-benzo[i,j]quinolizine-2-carboxylic acid and is disclosed in JP Patent No. 58,90,511 and U.S. Pat. No. 4,399,134. Nadifloxacin has an asymmetric carbon atom at the 5-position thereof. RS-(xc2x1)-Nadifloxacin comprises two optically active isomers. In describing an optically active compound, the prefixes R and S or D and L are used to denote the absolute configuration of the molecule about its chiral centre(s). The prefixes (+) and (xe2x88x92) or d and l are employed to designate the sign of rotation of plane-polarized light by the compound, with (xe2x88x92) or l meaning that the compound is levorotatory. A compound prefixed with (+) or d is dextrorotatory. Compounds having a single chiral centre exist as a pair of enantiomers, which are identical except that they are non-superimposable mirror images of one another. A one-to-one mixture of enantiomers is often referred to as a racemic mixture. Racemic RS-(xc2x1)-Nadifloxacin derives its biological activity primarily from the S-(xe2x88x92)-enantiomer. The optically active S-(xe2x88x92)-Nadifloxacin [xcex1]20D=xe2x88x92312.0 is obtained as disclosed in Chem. Pharm. Bull 44 (1996), page nos. 642-5 and Jpn. Kokai Tokyo Koho JP 63,192,753. The optically active R-(+)-Nadifloxacin, [xcex1]20D=+312.0, is obtained as disclosed in Jpn. Kokai Tokyo Koho JP 63,192,753. Pharmaceutical compositions of RS-(xc2x1)-Nadifloxacin are disclosed in U.S. Pat. No. 4,399,134 and U.S. Pat. No. 4,552,879. Although these cited patents disclose compositions of RS-(xc2x1)-Nadifloxacin for oral, parenteral and topical use, the only commercial product containing RS-(xc2x1)-Nadifloxacin as an active antibacterial compound is the commercial product named Acuatim(copyright). Acuatim(copyright) is available as a cream and a lotion and incorporates racemic RS-(xc2x1)-Nadifloxacin as 1% of its composition for the topical treatment of acne. Acuatim(copyright) has several drawbacks. It is intended only for topical use and is registered only for the treatment of acne caused by Propionibacterium species. One report has appeared on the in-vitro activity of the fluoroquinolone, Nadifloxacin, against methicillin resistant isolates of Staphylococcus aureus from patients with skin infections (see Nishijima et al., Drugs 49 (Suppl.) 230-232, 1995). There is no report of RS-(xc2x1)-Nadifloxacin being approved for systemic use against any microbial infections, whether for sensitive or resistant microbial strains.
S-(xe2x88x92)-Nadifloxacin is reported in Chem. Pharm, Bull 44 (1996) pages Nos. 421-423 to be approximtely twice as active in-vitro as racemic Nadifloxacin against Gram-positive and Gram-negative bacteria. There is no previous report, however, of the activity of S-(xe2x88x92)-Nadifloxacin in in-vivo systems against Gram-positive bacteria, Gram-negative bacteria, anaerobes, mycobacteria and emerging nosocomial pathogens.
RS-(xc2x1)-Nadifloxacin is reported to exist in two crystalline forms, one as an anhydrate and the other as a hemihydrate (M. Kido and K. Hashimoto, Chem. Pharm. Bull, 42, 872 (1994)). There is no previous report, however, of any hydrate forms of S-(xe2x88x92)-Nadifloxacin, although a non-hydrate form is reported (K. Hashimioto et al., Chem. Pharm. Bull., 44,642 (1996)).
There is no previous report of the utility of optically pure benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof of the invention in pharmaceutical compositions. There is also no previous report of the systemic or topical use of optically pure benzoquinolizine carboxylic acids, their derivatives, salts and hydrates thereof of the invention, either alone or in compositions for treatment of microbial infections diseases or disorders.
Our pending PCT patent application No. PCT/IN99/00016 filed on May 7, 1999 describes optically pure and racemic benzoquinolizine carboxylic acids, derivatives and salts thereof for treatment of infections caused by Mupirocin-resistant bacterial strains such as Mupirocin-resistant staphylococci, Mupirocin-resistant streptococci and other Mupirocin-resistant Gram-positive and Mupirocin-resistant Gram-negative bacteria, and for treatment of dermal diseases such as impetigo, folliculitis, infected dermatitis, wounds and bums. The subject matter of PCT application PCT/IN99/00016 is incorporated herein by reference.
The present inventors obtained optically pure isomers of Nadifloxacin and have conducted extensive studies to show that:
1. S-(xe2x88x92)-Nadifloxacin is found to exist not only as an anhydrate but also as three new different hydrates designated as S-(xe2x88x92)-Nadifloxacin.nH2O, wherein n is equal to 0.2, 0.5 or 0.75. Among these forms of the anhydrate and the three hydrates, S-Nadifloxacin.0.2 H2O is now specifically found to be preferred as a stable, non-hygroscopic crystalline modification which is distinguished by an increased stability, neither losing the water content therein nor absorbing moisture over a wide range of ambient relative humidity conditions. In contrast, moisture absorption by the anhydrate varies according to its method of preparation and the relative humidity conditions to which it is subjected. The other two hydrates, viz. the hemihydrate, S-(xe2x88x92)-Nadifloxacin.0.5H2O and the hemisesquihydrate, S-nadifloxacin.0.75H2O, both revert to S-nadifloxacin.0.2H2O when dried in vacuo at ambient temperature conditions.
S-(xe2x88x92)-Nadifloxacin.0.2H2O has considerable advantages over the anhydrate, the hemihydrate and the hemisesquihydrate in storage and handling and in the preparation of medicament forms. In particular, specially in tropical and subtropical climates, where the ambient humidity is usually greater than 70%, difficulty is encountered in the storage and handling of the anhydrate. In the preparation of pharmaceutical preparations, such as tablets, containing the anhydrate, the operations must be carried out with attention to absorption or desorption of water of crystallisation. More specifically, a room in which to handle the anhydrate must be kept at low humidity and conversely, a room in which to handle the hemihydrate and hemisesquihydrate must be kept at low temperatures and high humidity. Unless these conditions are provided, these compounds or preparations containing these compounds would change in weight, and thus would not serve for practical purposes and would lose their commercial value. By using a stable non-hygroscopic, free-flowing active compound, as is provided by S-(xe2x88x92)-Nadifloxacin 0.2 H2O, a satisfactory dosing consistency and accuracy is achieved during the preparation of medicaments, which increases safety and therefore minimizes the risk to the patient.
2. Crystalline salts of S-(xe2x88x92)-Nadifloxacin, especially sodium, potassium, and arginine salts, and hydrates thereof have been identified with increased aqueous solubility over S-(xe2x88x92)-Nadifloxacin, and consequently with superior properties for use in the preparation of parenteral formulations, and with advantages of improved oral bioavailability in solid oral dosage forms.
3. Derivatives of S-(xe2x88x92)-Nadifloxacin are identified at the sites of the 2-COOH fiuction and the 4xe2x80x2-hydroxy moiety of the 9-(4xe2x80x2-hydroxypiperidino) group respectively, and salts and hydrates thereof.
4. Processes are described to obtain the optically pure benzoquinolizine carboxylic acids of the invention, the derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof mentioned under items 1-3 above and as described in detail below.
5. RS-(xc2x1)-Nadifloxacin, S-(xe2x88x92)-Nadifloxacin and optically pure benzoquinolizine carboxylic acids, the derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof have high activity against Mupirocin-resistant microbial strains such as Mupirocin-resistant staphylococci, Methicillin-resistant staphylococcus aureus and Quinolone-resistant Staphylococcus aureus, coagulase negative staphylococci, such as Methicillin-resistant Staphylococcus epidermidis (MRSE), enterococci, betahemolytic streptococci and viridans group of streptococci.
6. RS-(xc2x1)-Nadifloxacin, S-(xe2x88x92)-Nadifloxacin and optically pure benzoquinolizine carboxylic acids, the derivatives, salts, pseudopolymorphs, polyrnorphs and hydrates thereof have activity against mycobacteria and newly emerging nosocomial pathogens such as Chryseobacterium meningosepticum. 
7. S-(xe2x88x92)-Nadifloxacin and optically pure benzoquinolizine carboxylic acids, the derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof have 2-4 times higher antimicrobial activity than racemic-Nadifloxacin against Mupirocin-resistant staphylococci, Methicillin-resistant Staphylococcus aureus (MRSA), Quinolone-resistant Staphylococcus aureus, coagulase negative staphylococci, such as Methicillin-resistant Staphylococcus epidermidis (MRSE), enterococci, betahemolytic streptococci and viridans group of streptococci, mycobacteria and newly emerging nosocomial pathogens such as Chryseobacterium meningosepticum. 
8. S-(xe2x88x92)-Nadifloxacin and optically pure benzoquinolizine carboxylic acids, the derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof are not only bacteriostatic but also bactericidal towards Mupirocin-resistant staphylococci, Methicillin-resistant Staphylococcus aureus (MRSA), Quinolone-resistant Staphylococcus aureus, coagulase negative staphylococci, such as Methicillin-resistant Staphylococcus epidermidis (MRSE), enterococci at concentrations 2-4 times lower than that of RS-(xc2x1)-Nadifloxacin.
9. S-(xe2x88x92)-Nadifloxacin and optically pure benzoquinolizine carboxylic acids, the derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof have 2-4 times higher antimicrobial activity against Gram-positive pathogens such as staphylococci and enterococci and Gram-negative pathogens such as E.coli, Klebsiella, Proteus, Serratia and Citrobacter in the acidic environments encountered in infection such as urinary tract infections.
10. S-(xe2x88x92)-Nadifloxacin, its derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof have high potency against efflux pump-bearing Staphylococcus strains and are thus of unique value in treating infections caused by antibiotic-resistant microorganisms for which the resistance mechanism is due to the presence of efflux pumps.
11. S-(xe2x88x92)-Nadifloxacin, its derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof have high propensity to display resistance to resistance development, which has been shown in studies involving sequential transfers/passages of a S.aureus strain through respective drug containing media.
12. The acute intravenous toxicity of S-(xe2x88x92)-Nadifloxacin and its arginine salt is significantly lower than RS-(xc2x1)-Nadifloxacin (Biological Example 3).
13. S-(xe2x88x92)-Nadifloxacin, its derivatives, salts and hydrates thereof have a favourable toxicity profile in comparison with other fluoroquinolone drugs in clinical use in respect of cytotoxic effect on various cell lines (Biological Example 4), phototoxicity (vide infra) and cardiotoxicity.
14. The oral bioavailability of S-(xe2x88x92)-Nadifloxacin is 2-times higher than that of RS-(xc2x1)-Nadifloxacin (Biological Example 5).
Through their extensive studies, the present inventors have shown for the first time a novel expanded set of clinically desired antimicrobial attributes of efficacy and safety of S-(xe2x88x92)-Nadifloxacin, which have been not reported in the literature since the first disclosure of S-(xe2x88x92)-Nadifloxacin in JP 63,192,753 about twelve years ago and of RS-(xc2x1)-Nadifloxacin in JP 58,90,511 about twenty years ago. New hydrates, salts, derivatives, pseudopolyrnorphs, polymorphs and compositions of S-(xe2x88x92)-Nadifloxacin have also been identified by the present inventors, which have in addition to their biological properties mentioned above, newer physico-chemical properties, thus permitting their utility in a clinical and commercial exploitation in newer compositions for newer diseases and newer methods of systemic and topical treatment that were hitherto not possible.
It is, thus, an object of the present invention to provide S-(xe2x88x92)-optically pure benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polyrnorphs and hydrates thereof, of the formula I, substantially free of their R-(+)-isomers. 
It is another object of the present invention to provide a process or processes for preparing the novel optically pure S-(xe2x88x92)-optically pure benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof, of the Formula I.
A further object is to provide pharmaceutical compositions comprising optically pure S-(xe2x88x92)-benzoquinolizine carboxylic acids, the derivatives, pseudopolymorphs, hydrates and salts thereof as potent antibacterial agents for treating systemic and topical bacterial infections, especially infections caused by resistant Gram-positive, sensitive and resistant Gram-negative organisms, mycobacterial infections and nosocomial pathogen infections while avoiding the toxic effects associated with the administration of their R-(+)-isomers.
Another object of this inventions relates to a method of treatment of infections caused by Mupirocin-resistant bacterial strains such as Mupirocin-resistant staphylococci, Mupirocin-resistant streptococci and other Mupirocin-resistant Gram-positive and Mupirocin-resistant Gram-negative bacteria, and of dermal diseases and disorders such as impetigo, folliculitis, infected dermatitis, wounds and burns. Treatment comprises oral, parenteral, administration and/or topical application of an effective amount of a composition of S-(xe2x88x92)-Nadifloxacin or optically pure benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof of formula I, or of a composition of RS-(xc2x1)-Nadifloxacin and pharmaceutically acceptable salts thereof.
A further object of the invention includes methods for treating the infections in humans and animals caused by Gram-positive, Gram-negative and anaerobic bacteria, resistant Gram-positive organism such as Methicillin-resistant Staphylococcus aureus (MRSA), Quinolone-resistant Staphylococcus aureus, coagulase negative staphylococci, such as Methicillin-resistant Staphylococcus epidermidis (MRSE), enterococci, betahemolytic streptococci and viridans group of streptococci, mycobacteria and newly emerging nosocomial pathogens such as Chryseobacterium meningosepticum, and Gram-negative pathogens such as E. coli, Klebsiella, Proteus, Serratia Citrobacter and Pseudomonas, while avoiding the toxic effects that are associated with the racemic mixture of Nadifloxacin by administering systemically or topically S-(xe2x88x92)-Nadifloxacin or optically pure S-(xe2x88x92)-benzoquinolizine carboxylic acids, their derivatives, salts, pseudopolymorphs, polymorphs and hydrates thereof to the affected human or animal.